ENZYMATIC METHYLATION OF ARSENIC COMPOUNDS - ASSAY, PARTIAL-PURIFICATION, AND PROPERTIES OF ARSENITE METHYLTRANSFERASE AND MONOMETHYLARSONIC ACID METHYLTRANSFERASE OF RABBIT LIVER

A rapid, accurate, in vitro assay utilizing radioactive S-adenosylmeth ionine (SAM) has been developed for the methylation of arsenite and mo nomethylarsonate (MMA) by rabbit liver methyltransferases. The assay h as been validated by separating, identifying, and measuring the produc ts of the reaction using chloroform extraction, ion exchange chromatog raphy, TLC, or HPLC. The enzymes involved in this pathway, arsenite me thyltransferase and MMA methyltransferase, have been purified approxim ately 2000-fold from rabbit liver. After gel electrophoresis, a single band is obtained with both enzyme activities in it. The pH optima for purified arsenite methyltransferase and monomethylarsonic acid methyl transferase are 8.2 and 8.0, respectively. A thiol, S-adenosylmetkioni ne, and arsenite are required for the partially purified arsenite meth yltransferase that catalyzes the synthesis of monomethylarsonate. A di fferent enzyme activity that catalyzes the methylation of monomethylar sonate to dimethylarsinate also requires SAM and a thiol. Even though arsenite methyltransferase and monomethylarsonate methyltransferase ha ve different substrates, pH optima, and saturation concentrations for their substrates, whether the two activities are present on one protei n molecule or different protein molecules is still uncertain. Both act ivities have a molecular mass of 60 kDa as determined by gel exclusion chromatography. There is no evidence at the present time for these en zyme activities being on different protein molecules. Neither arsenate , selenate, selenite, or selenide are methylated by the purified enzym e preparations. Results from the use of crude extracts, often called c ytosol, to study the properties of these methyltransferases dealing wi th arsenic species should be viewed with caution since such crude extr acts contain inhibiting and other interfering activities.